Peptide variants of protein A

Abstract

Z domain variants of staphylococcal protein A have significantly reduced size but possess IgG-binding affinity equivalent to the wild type Z domain. These Z domain variants are suitable for use in affinity chromatography purification of proteins and in the treatment of staphylococcic diseases.

We claim:  
     
       1. A compound represented by Formula (II):                    
       where 
       X 1  is selected from the group consisting of H, C 1 -C 6 alkanoyl, and Z-Ala-Val-AA 3 -AA 4 -AA 5  (SEQ ID NO:2);  
       where  
       Z is selected from the group consisting of H and C 1 -C 6 alkanoyl;  
       AA 3  is selected from the group consisting of Asp, Arg, and Ala;  
       AA 4  is selected from the group consisting of Asn and Gln; and  
       AA 5 is selected from the group consisting of Lys, Gly, and Ser;  
       AA 6  is selected from the group consisting of Phe and Gly;  
       AA 7  is selected from the group consisting of Asn and Trp;  
       AA 8  is selected from the group consisting of Lys and Met;  
       AA 9  is selected from the group consisting of Glu, Gln, and Arg;  
       AA 12  is selected from the group consisting of Asn, Ala, and Arg;  
       AA 13  is selected from the group consisting of Ala and Arg;  
       AA 33  is selected from th e group consisting of Gln and Lys;  
       AA 36  is selected from the group consisting of Lys and Arg; and  
       X 2  is selected from the group consisting of OR 1  and NR 1 R 2  where R 1  and R 2  are independently selected from the group consisting of H, C 1 -C 6 alkyl, C 6 -C 12 aryl and C 6 -C 12 aryl-C 1 -C 6 alkyl;  
       which compound is covalently linked to a macromolecule to form a conjugate. 
     
     
       2. The conjugate of claim  1  wherein the macromolecule is a solid support. 
     
     
       3. The conjugate of claim  1  wherein the compound is selected from the group consisting of:                                      
       where 
       X 1  is selected from the group consisting of H and C 1 -C 6 alkanoyl;  
       Z is selected from the group consisting of H and C 1 -C 6 alkanoyl; and  
       X 2  is selected from the group consisting of OR 1  and NR 1 R 2  where R 1  and R 2  are independently selected from the group consisting of H, C 1 -C 6 alkyl, C 6 -C 12 aryl and C 6 -C 12 aryl-C 1 -C 6 alkyl.  
     
     
       4. The conjugate of claim  1  wherein the compound is selected from the group consisting of:                    
       where 
       X 1  is selected from the group consisting of H and C 1 -C 6 alkanoyl;  
       Z is selected from the group consisting of H and C 1 -C 6 alkanoyl; and  
       X 2  is selected from the group consisting of OR 1  and NR 1 R 2  where R 1  and R 2  are independently selected from the group consisting of H, C 1 -C 6 alkyl, C 6 -C 12 aryl and C 6 -C 12 aryl-C 1 -C 6 alkyl.  
     
     
       5. A compound represented by Formula (II):                    
       where 
       X 1  is selected from the group consisting of H, C 1 -C 6 alkanoyl, and Z-Ala-Val-AA 3 -AA 4 -AA 5  (SEQ ID NO:2);  
       where  
       Z is selected from the group consisting of H and C 1 -C 6 alkanoyl;  
       AA 3  is selected from the group consisting of Asp, Arg, and Ala;  
       AA 4  is selected from the group consisting of Asn and Gln; and  
       AA 5  is selected from the group consisting of Lys, Gly, and Ser;  
       AA 6  is selected from the group consisting of Phe and Gly;  
       AA 7  is selected from the group consisting of Asn and Trp;  
       AA 8  is selected from the group consisting of Lys and Met;  
       AA 9  is selected from the group consisting of Glu, Gln, and Arg;  
       AA 12  is selected from the group consisting of Asn, Ala, and Arg;  
       AA 13  is selected from the group consisting of Ala and Arg;  
       AA 33  is selected from the group consisting of Gln and Lys;  
       AA 36  is selected from the group consisting of Lys and Arg; and  
       X 2  is selected from the group consisting of OR 1  and NR 1 R 2  where R 1  and R 2  are independently selected from the group consisting of H, C 1 -C 6 alkyl, C 6 -C 12 aryl and C 6 -C 12 aryl-C 1 -C 6 alkyl;  
       which compound is fused to a selected polypeptide to form a fusion protein. 
     
     
       6. The fusion protein of claim  5  wherein the compound of Formula (II) is specifically cleavable from the amino acid sequence of the selected polypeptide. 
     
     
       7. The fusion protein of claim  5  wherein the compound is selected from the group consisting of:                                      
       where 
       X 1  is selected from the group consisting of H and C 1 -C 6 alkanoyl;  
       Z is selected from the group consisting of H and C 1 -C 6 alkanoyl; and  
       X 2  is selected from the group consisting of OR 1  and NR 1 R 2  where R 1  and R 2  are independently selected from the group consisting of H, C 1 -C 6 alkyl, C 6 -C 12 aryl and C 6 -C 12 aryl-C 1 -C 6 alkyl.  
     
     
       8. The fusion protein of claim  5  wherein the compound is selected from the group consisting of:                    
       where 
       X 1  is selected from the group consisting of H and C 1 -C 6 alkanoyl;  
       Z is selected from the group consisting of H and C 1 -C 6 alkanoyl; and  
       X 2  is selected from the group consisting of OR 1  and NR 1 R 2  where R 1  and R 2  are independently selected from the group consisting of H, C 1 -C 6 alkyl, C 6 -C 12 aryl and C 6 -C 12 aryl-C 1 -C 6 alkyl.